Most of the existing information about DNA replication in mammalian cells comes from studies of lytic viruses. Some aspects of the regulation of DNA replication cannot be studied readily in these systems, because most lytic viruses lack a clearly defined replication control. BPV is maintained as a stable replicating episome in transformed rodent cells. Thus, study of bovine papillomavirus (BPV) DNA replication offers a model for copy number control and segregation in mammalian cells. The overall goals of the proposed research are to understand the properties of DNA replication of BPV. These goals will be addressed by studying viral replication at two different levels. First, a detailed study of the viral cis-acting elements and the two virus encoded transacting factors that are involved in replication will be undertaken. This will serve to define the role of the factors and elements in the replication process. This approach will include the establishment of an in vitro replication system for BPV. Second, in vivo studies will be initiated to determine how BPV can establish and maintain itself as a stable plasmid. This question will be addressed both by detailed kinetic analysis of replication in vivo, and by determination of the requirements for viral genes and viral gene regulation in copy number control and maintenance. The experimental approach will be to uncouple the expression of the trans-acting factors required for replication, from viral transcriptional control. This will be accomplished by generation of stable cell lines expressing the required trans-acting factors from heterologous promoters. These cell lines will be used to determine to what extent viral gene regulation is responsible for the replication properties of BPV. Finally, BPV is firmly established as the prototype papillomavirus, serving as a model for replication, gene regulation and transformation. The human papillomaviruses, which are widely considered to be involved in the development of a variety of malignancies, most notably cervical carcinomas, are difficult to study at the molecular level. Although very little is know about the replication properties of the human viruses, it is likely that study of BPV replication will furnish insight into the life cycle of the human viruses.